Biological specimen processing and embedding apparatus

ABSTRACT

A COMBINATION CAPSULE AND MOLD APPARATUS AND TREATMENT OF A BIOLOGICAL SPECIMEN BY THE USE THEREOF IS DESCRIBED WHEREIN A BIOLOGICAL SPECIMEN CAN BE PLACED INSIDE A CAPSULE COMPRISING AN OPEN-TOPPED BOX-LIKE OPEN MOLD HAVING A PERFORATED BOTTOM WALL AND REMOVABLE PERFORATED COVER. AFTER THE SPECIMEN HAS BEEN PROCESSED INSIDE THE CAPSULE WITH DESIRED FLUIDS, THE COVER IS REMOVED FROM THE CAPSULE TO FORM AN OPEN MOLD, THE SPECIMEN IS TRANSFERRED FROM THE OPEN MOLD TO AN OPEN-TOPPED BOX-LIKE BASE MOLD, AND THE OPEN MOLD IS PLACED IN COUPLED RELATION OVER THE BASE MOLD. MOLTEN PARAFFIN IS THEN   POURED THROUGH THE OPEN MOLD INTO THE BASE MOLD. AFTER THE PARAFFIN HARDENS, THE OPEN MOLD AND THE RIGIDLY ATTACHED PARAFFIN BODY CONTAINING THE SPECIMEN ARE SEPARATED FROM THE BASE MOLD, AND THE OPEN MOLD IS MOUNTED IN THE HOLDER OF A MICROTOME FOR SUBSEQUENT SLICING OF THE PARAFFIN-EMBEDDED SPECIMEN.

Sept. 17, 1974 J, B, MGcoRMlcK BIOLDGICAL SPECIMEN PROCESSING ANDEMBEDDING APPARATUS Original Filed )lay 1, 1968 F|6URE 2. 488

FIGURE 3.

FIGURE 4.

FIGURE 5.

FIGURE 6.

r/zFIGURl- 7.

INVENTOR JAMES BENJAMIN MC CORMICK ATTORNEY United States Patent OfliceRe. 28,165 Reissued Sept. 17, 1974 28,165 BIOLOGICAL SPECIMEN PROCESSINGAND EMBEDDING APPARATUS James Benjamin McCormick, Hinsdale, lll.,assignor to Miles Laboratories, Inc., Elkhart, Intl` Original No.3,674,396, dated July 4, 1972, Ser. No.

74,159, Sept. 21, 1970, which is a continuation of abandoned applicationSer. No. 725,722, May 1, 1968. Ap-

plication for reissue May 21, 1973, Ser. No. 361,973

Int. Cl. B29c 1/00I I/02, 1/14 U.S. Cl. 425-117 3 Claims Matter enclosedin heavy brackets appears in the original patent but forms no part ofthis reissue specification; matter printed in italics indicates theadditions made by reissue.

ABSTRACT OF THE DISCLOSURE A combination capsule and mold apparatus andtreatment of a biological specimen by the use thereof is de scribedwherein a biological specimen can be placed inside a capsule comprisingan open-topped box-like open mold having a perforated bottom wall and aremovable perforated cover. After the specimen has been processed insidethe capsule with desired fluids, the cover is removed from the capsuleto form an open mold. the specimen is transferred from the open mold toan open-topped box-like base mold, and the open mold is placed incoupled relation over the base mold. Molten paraffin is then pouredthrough the open mold into the base mold. After the paraffin hardens,the open mold and the rigidly attached paraffin body containing thespecimen are separated from the base mold, and the open mold is mountedin the holder of a microtome for subsequent slicing of theparafiin-embedded specimen.

This application is a continuation of application Ser. No. 725,722,filed on May 1. 1968, now abandoned.

BACKGROUND AND PRIOR ART It is well known in the art that biologicaltissues can be sliced into thin sections on a microtome for subsequentmicroscopic examination by a pathologist, for example. In order toobtain a satisfactory thin section wherein the tissue is not distorted,the tissue must be substantially rigid at the time of microtome slicing.One commonly employed procedure of providing this rigidity is to embedthe tissue in hardened paraffin.

In order to properly use the paraffin. the tissue must first beprocessed `with several iluids to dehydrate the tissue, to clear thetissue with a suitable oil and to iniiltrate the tissue with a paraffinwax or a combination of wax and resinous material. This processing hasbeen conveniently carried out by placing the specimen in afluid-permeable capsule and successively submerging the capsule in thenecessary Fluids. The resulting processed specimen is then removed fromthe capsule and embedded in a block of paratln wax for subsequentmounting in a microtome for slicing. The capsule can be reused forprocessing specimens.

Various apparatus have been employed in the prior art for embedding theIspecimens in parafiin. Most of such apparatus were crude and resulted inembedded specimens having many disadvantages. The apparatus of US. Pat.No. 2,996,762 provided a substantial improvement in embedding apparatus.This improvement comprised an embedding box which resulted in a unitarystructure of paraffin block and box designed for easv and accurateclamping in a holder on a microtome. While the apparatus of U.S. Pat.No. 2,996,762 had many advantages, it

still required a separate capsule apparatus for processing the specimenprior to the paraffin embedding szep.

OBJECTS AND SUMMARY OF THE INVENTION It is an object ot" the presentinvention to provide improved apparatus for embedding a specimen inparaffin which can also be used to mount the specimen in a microtome.

It is another object of the present invention to provide combinationapparatus which can be used both to process the specimen prior toembedding and to embed the specimen in paratiin.

In accordance with the present invention, a combination capsule and moldapparatus useful in the procesing, paraffin embedding` and slicing ofbiological specimens is provided which comprises an open-topped boxlikebase mold having a bottom wall, an open-topped boxlike open mold membercapable of placement in coup'ed relation on the open top of said basemold, said open mold having a perforated bottom wall. and a removablecover for said open mold` said open mold with the removable cover inplace providing a fluid-permeable capsule in which a biological specimencan be treated with selected fluid, said base mold being capable ofaccom- Inodating a treated specimen transferred thereto from saidcapsule. placement of said molds in said coupled relation while aspecimen is disposed within said base mold permitting said base mold tobe Filled with molten parafiin poured ther-cinto through said open moldwith said cover removed until said paraflin extends above the perforatedbottom wall of said open mold. said base mold defining the shape of theparaffin body when hardened having a specimen-embedded face along thebottom wall of the base mold. and the perforated bottom wall of saidopen mold affording means for anchoring said paraffin body to said openmold such that on removal of said base mold from said coupled relationsaid open mold and parafln body remain as a finished unit capable ofbeing mounted in a specimen holder of a microtome.

DESCRIPTION OF THF DRAWINGS FIG. l is a perspective view of the separateparts of the combination capsule and mold apparatus in explodedrelation;

FIG. 2 is a cross-sectional `view taken generally along plane A-A ofFIG. l showing the capsule formed from the cover and open mold member,said capsule containing a biological specimen;

FIG. 3 is a cross-sectional view taken generally along plane A A of FIG,l showing the open mold member with the cover removed and said open moldmember fitted over the base mold` said base mold containing a biologicalspecimen;

FIG. 4 is a similar view to FIG. 3 showing the onen mold member and basemold containing embeddng paraffin;

FIG. 5 is a similar view to FIG. 4 with the base mold removed;

FIG. 6 is a similar view to FIG. 3 showing the use of a base mold havinga smaller storage area to accommodate a smaller specimen as shown: and

FIG. 7 is a side view in partial cross-section showing the unitarystructure of FIG. 5 supported in a microtome specimen holder.

DESCRIPTION OF THE INVENTION Referring to FIGS. l to 3, the apparatus ofthe Present invention comprises a base mold Il), a multi-purpose capsuleand open mold member 12. and a removable cover 14. Base mold I() isfo'med in a generallv openformed box-like rectangular' shape with asubstantially fiat bottom wall 16, transverse sidewalls I8 and 19,

longitudinal sidewalls 20 and 2l and an open top 22. The Lipperpositions of the sidewalls 18 to 21 are offset outwardly to form anannular horizontal surface or ledge 24 within the open top 22 which issubstantially parallel to bottom wall 16, Longitudinal lip walls 23 and25 extend upwardly and horizontally outwardly from the upper edges ofthe longitudinal sidewalls 20 and 21 and connect said sidewalls withupstanding longitudinal tins 26 and 27 respectively. Transverse lipswalls 30 and 31 extend upwardly and horizontally outwardly from theupper edges of the transverse sidewalls 18 and 19 and connect saidsidewalls willi downturned transverse fins 28 and 29 respectively. Thislatter feature is best shown in FlG. 3. Base mold 10 is preferablyformer from metal or other suitable material having a relatively highcoefhcient of heat transmission.

Referring to FlGS. l, 2 and 3, the multi-purpose capsule and open moldmember 12 is formed in a generally open-topped box-like rectangularshape with a bottom wall 32 having a plurality of perfor-ations 34,longitudinal sidewalls 36 and 37, transverse sidewalls 38 and 39, and anopen top 40. Sidewalls 36, 37, 38 and 39 have coplanar upper edgesurfaces 42, 43, 44 and 45, respectively. `which are normal to saidsidewalls and which form a substantially fiat annular end surfacedefining the open top 40.

The outer face of the transverse sidewall 38 is formed along its upperedge with an outwardly projecting ridge 48. Connected to and extendingdownwardly and outwardly from the upper edge portion of the transversesidewall 39 is a slanted wall 58, and the adjacent end portions of thelongitudinal sidewalls 36 and 37 extend outwardly beyond the transversesidewall 39 to join the slanted wall 58 along slant edges 52 and 54respectively. The upper edge portions of the transverse sidewall 39 andof the slanted wall 58 are cut away as at 64 to form a transverse slot66 which affords access to the transverse chamber 63 of generallytriangular crosssection which is formed between the outer face ofsidewall 39 and the underside of the slanted wall 58. Capsule member 12is preferably formed from an organo-plastic or other suitable materialhaving a lower coefficient of heat transmission than that of the basemold 10.

Referring to FIG. 1, removable cover 14 is formed of a generallyrectangular fiat plate 68 which prefer'- ably has a plurality ofperforations 70 therein. Plate 68 has longitudinal edges 72 and 74 andtransverse edges 76 and 78. A tongue portion 8() extends downwardly andoutwardly from the mid portion of the edge 78 and a tab or handleportion 82 extends from the mid portion of the edge 76 coplanar with theplate 68. At each side of the tab portion 82 a spring latch member 84having a generally S-shaped cross-section, as viewed in FIGS. 1 and 2,is attached to edge 76 and extends downwardly in a direction generallynormal to plate 68. Cover 14 is preferably formed from metal, but othersuitable materials, stlch as organo-plastics, can also be used.

ln order to utilize this novel apparatus, a biological specimen 86 isplaced within the member 12 as shown in FIG. 2. The cover 14 is thenfitted over the open top 4U of member l2 by inserting the tongue 80through the slot 66 and snapping the latch members 84 over ridge 48 asshown in FlG. 2 to close the capsule. The crosssectional dimensions ofthe perforations 34 in the bottom wall 32 and of the pcrforations 70 inthe cover plate 68 are both smaller than any corresponding dimension ofthe specimen 86, so that the specimen will be retained within the soformed capsule. The fluid-permeable capsule containing the specimen canthen be successively placed in solutions of dehydrating agents, clearingagents and parafiin wax to properly prepare the specimen for subsequentembedding in paraffin. The processing tiuids easily pass through theperforations 34 and 70.

The cover 14 is then removed from the capsule member 12 by lifting up onhandle 82. Specimen 86 is then removed from member 12 and placed in thebase mold 10 as shown in FIG. 3. The specimen is oriented so that theportion of the specimen to be ultimately sliced first in a microtome isin Contact with the bottom wall 16. A small amount of molten paraffincan then be poured into the base mold 10 to harden around specimen 86and maintain the desired orientation thereof with respect t0 the bottomwall 16.

The open mold member 12 is then placed on the base mold 10 in nested orCoupled relation therewith as shown in FIG. 3. ln this position themember 12 rests on the annular horizontal surface 24 with the outersurfaces of the sidewalls 36, 37 and 38 of said member 12 substantiallyin abutment with the inner surfaces of the lip walls 25, 23 and 30respectively. The lower edge 60 of slanted wall 58 is substantially inabutment with lip wall 31. The member 12 is thus retained in the properposition. The flat annular Lipper surface of mold member 12 formed bycoplanar Lipper edge surfaces 42, 43, 44 and 45 is then substantiallyparallel to the bottom wall 16 of the base mold 10.

Hot molten paraffin is then poured through perforations 34 of the openmold member 12 into base mold 10 until the base mold 10 andsubstantially all of the mold member 12 are filled with paraffin. Sincethe base mold preferably has a coefficient of heat transmission greaterthan that of the open mold member, heat from the paraffin is dissipatedthrough the bottom and side walls of the base mold 10 more rapidly thanthrough the sidewalls of the open mold member 12 with the result thatthe paraffin initially solidifies against the inner surfaces of the basemold 10. The shrinkage of the parafn when cooling also moves toward thebase mold. When the entire paraffin body 88 has hardened, it has thestructure shown in cross-section in FIG. 4 including an upper shrinkagemeniscus 90. It will be observed that the perforated bottom wall 32 ofthe open mold member 12 is embedded in the solidified paraffin body 88so that the open mold member 12 is rigidly anchored to the finishedparafiin body 88 to form a finished unit. The bottom and sidewalls ofthe base mold 10 serve to define the shape of the finished paraffin body88 when hardened with the specimen oriented along the base mold bottomwall 16. The apparatus of the present invention requires about 20-30weight percent less paraffin to embed a given size specimen than theapparatus of U.S. Pat. No. 2,996,762.

The fins 26, 27, 28 and 29 extending from the periphery of the open topof the base mold 1t) aid in conducting heat away from the contactinterfaces between the open mold member 12 and the base mold 10.

Base mold 10 can then be easily removed from the finished paraflin body88 to form the finished unit shown in cross-section in FIG. 5 having aspecimen embedded face 92. The upper surface of member 12 is parallelwith face 92. This finished unit can then be easily mounted in thespecimen holder of a microtome as shown in partial cross-section in FIG.7. The specimen holder 94, which is either an integral part of themicrotome or is supported by the microtome, has a substantially fiataligning face 96 which is in a plane parallel to the path of travel 98of a microtome blade 100 which has a reciprocal motion with respect tothe specimen holder. The specimen holder 94 also has a stationarygripping means 102 and a movable gripping means 104 both having grippingfaces normal to face 96.

The mold member portion of the finished unit is placed into the specimenholder 94 with the annular end surface of the open mold member, definedby edge surfaces 42 to 4S, seated against the fiat aligning face 96 ofholder 94. Sidewall 36 is placed in contact with stationary grippingmeans 192 and movable gripping means 104 is suitably locked intogripping contact with sidewall 37. The finished unit is thus rigidlysupported in holder 94 with the specimen-embedded face 92 thereofdisposed in a plane parallel with the path of travel 98 of the microtomeblade 100. This physical relationship between the specimen-embedded face92 and the blade motion along path 98 is desirable in order to obtainspecimen slices of uniform thickness. In practice the microtome holder94 would be moved to the left, as viewed in FIG. 7, until the face 92was over blade 100. The blade 100 would then be moved upward, as viewedin FIG. 7, relative to paraffin body 88 to cut a slice from thespecimen-embedded in body 88.

After the desired slice or slices have been cut from the specimen, thefinished unit can be removed from the holder 94 for storage. Since theannular end face of the open mold member 12 is parallel to the face 92of the specimenembedded paraffin body 88, whenever additional slices aresubsequently to be made, the repositioning of the member l2 against thealigning face 96 of the holder 94 will again place the face 92 in aplane parallel to the microtome blade motion.

In a laboratory handling a large number of specimens, it is necessarythat proper specimen identification be maintained. The apparatus of thepresent invention employs slanted wall 58 for this purpose. Wall 58 hasa surface that can be easily written upon with pencil or pen for theapplication of an identification designation. This enables the specimento be continuously identified through processing, embedding and slicing.

The base mold shown in FIGS. 1, 3 and 4 is intended primarily forspecimens having a fairly large size. If this particular base mold wereused with a fairly small specimen, an excessive amount of paraffin wouldbe needed to embed the specimen. The base mold l0' shown in crosssectionin FIG. 6 has a relatively small volume chamber formed by bottom wall 16and sidewalls, such as 18' and 19', which has a size not substantiallylarger than necessary to accommodate the small specimen 86'. Base moldshaving a variety of chamber sizes can be used in the apparatus of thisinvention. Except for the chamber sizes, however, the dimensions of thebase molds l0 of FIG. 6 are the same as those for the base mold 10 inFIGS. 1, 3 and 4 so that the same sized open mold member 12 can beemployed therewith. Since the finished mold-paraffin units are generallystored for future reference after specimen slices are made, it ispreferable to employ open mold members 12 having the same size for allspecimens, because this simplifies the storage and retrieval of theunits.

In summary, this invention relates to an improved apparatus combinationthat can be used to process biological specimens, embed such specimensin paraffin and support the embedded specimens in a microtome forsubsequent slicing.

I claim:

1. Combination capsule and mold apparatus useful in the processing,parafiin embedding, and slicing of biological specimens which comprises(l) an open-topped rectangular shaped base mold having four sidewallsand a flat imperforate bottom wall, each of said sidewalls having anextending fin wherein two of the opposing sidewalls have fins extendingin an upward direction from the open top of the base mold and the tworemaining opposing sidewalls have fins extending in a downward directionfrom the open top of the base mold, (2) an open-topped rectangularshaped open mold member having a perforated bottom wall, first andsecond opposing transverse sidewalls, third and fourth opposinglongitudinal sidewalls, said transverse and longitudinal sidewallshaving coplanar fiat upper surfaces, said open mold member having aslanted wall `with an exterior surface capable of being easily writtenupon extending downwardly and outwardly from the fiat upper surface ofthe first transverse sidewall, the third and fourth longitudinalsidewalls extending beyond said tirst transverse sidewall to join saidslanted wall to form a transverse chamber between said first transversesidewall, said slanted wall and said extensions of said third and fourthlongitudinal sidewalls, said open mold member also having a transverseslot located along the junction between the slanted wall and the fiat[supper] upper surface of the first transverse sidewall, said transverseslot providing upper access to said transverse chamber, said open moldmember further having an outwardly projecting ridge along the outersurface of the second transverse sidewall and communicating with theupper at surface of said sidewall, and (3) a removable cover for saidopen mold member, said removable cover formed of a perforatedrectangular flat plate with a tongue portion extending downwardly andoutwardly from the mid portion of one transverse edge of said plate, atab extending from the mid portion of the opposing transverse edge ofsaid plate coplanar with said plate, and a spring latch member locatedat the side of said tab and extending downwardly from said transverseedge of said plate, said cover capable of being removably attached tosaid open mold member with the tongue portion of said cover insertedthrough the transverse slot of said open mold member and the springlatch member of said cover placed over the outwardly projecting ridge ofsaid open mold member, said open mold member with the removable cover inplace and being free of the base mold providing a Huid-permeable capsulein which a biological specimen can be treated with selected uid, saidbottom wall of the open mold member and said removable cover both havinga plurality of perforations the cross-sectional dimensions of which aresmaller than any corresponding dimensions of a specimen to be processedin said apparatus, said base mold being capable of accommodating atreated specimen transferred thereto from said capsule, said open moldmember with said cover removed being capable of placement in coupledrelation on the top of said base mold while a specimen is disposedwithin said base mold wherein said coplanar fiat upper surfaces of saidopen mold member are parallel with said fiat bottom wall of said basemold, said coupled relation permitting said base mold to be filled withmolten paraffin poured thereinto through said open mold member with saidcover removed until said parafiin extends above the perforated bottomwall of said open mold member, said base mold defining the shape of theparaffin body when hardened having a specimen-embedded face along thebottom wall of the base mold and the perforated bottom wall of said openmold member affording means for anchoring said paraffin body to saidopen mold member such that on removal of said base mold from saidcoupled relation said open mold member and paraffin body remain as alinished unit capable of being mounted in a specimen holder for amicrotome, said coplanar flat upper surfaces of said open rnold membercapable of being employed as alignment means for properly positioningsaid finished unit in said microtome.

2. Combination capsule and mold apparatus useful in the processing,parajn embedding, and slicing of biological specimens which comprises,(1) an open-topped' rectangular shaped base mold having four sidewallsand a flat imperforate bottom wall, each of said sidewalls having andextending )in wherein two of the opposing sidewalls have )ins extendingin an upward direction from the open top of the base mold and the tworemaining opposing sidewalls have fins extending in a downward directionfrom the open top of the base mold, (2) an open-topped rectangularshaped open mold member having n perforated bottom wall, first andsecond opposing transverse sidewalls, third and fourth opposinglongitudinal sidewalls, said transverse and longitudinal sidewallshaving coplanar flat upper surfaces, said open mold member having aslanted wall with an exterior surface capable of being easily writtenupon extending downwardly and outwardly from the flat upper surface ofthe first transverse sidewall, the third and fourth longitudinalsidewalls e.\- tending beyond said first transverse sidewall to joinsaid slanted wall to form a transverse chamber between .mid jrsttransverse sidewall, said slanted wall and srt/'d extensions of saidthird und fourth longitudinal sidewalls,

said open mold member also having a transverse slot located along tlzejunction between the slanted wall and the flat upper surface of thefirst transverse sidewall, said transverse slot providing upper accessto said transverse chamber, and (3 a removable cover for said open moldmember, said removable cover formed of a perforated rectangular fiatplate with a tongue portion extending downwardly from said plate, saidcover capable 0f being removably attached t0 said open mold member withthe tongue portion of said cover mating against said open mold member,said open mold member with the removable cover in place and being freeof the base mold providing a Huid-permeable capsule in which abiological specimen can be treated with selected fluid, said bottom wallof the open mold member and said removable cover both having a pluralityof performations the cross-sectional dimensions of which are smallerthan any corresponding dimensions of a specimen to be processed in saidapparatus, said base mold being capable of accommodating a treatedspecimen transferred thereto from said Capsule, said open mold memberwith said cover removed being capable of placement in coupled relationon the top of said base mold while a specimen is disposed within saidbase mold wherein said coplanar flat upper surfaces of said open moldmember are parallel with said fiat bottom wall of said base mold, saidcoupled relation permitting said base mold to be lled with moltenparaffin poured thereinto through said open mold member with said coverremoved until said parafjin extends above the perforated bottom wall ofsaid open mold member, said base mold defining the shape of the para/finbody when hardened having a specimen-embedded face along the bottom wallof the base mold and the perforated bottom wall of said open mold memberaffording means for anchoring said parain body to said open mold membersuch that on removal of said base mold from said coupled relation saidopen mold member and paraffin body remain as a finished unit capable ofbeing mounted in a specimen holder for a microtome, said coplanar flatupper surfaces of said open mold member capable of being employed asalignment means for properly positioning said finished unit in saidmicrotome.

3. Apparatus according to Claim 2 wherein said cover is capable of beingremovably attached to said open mold member with the tongue portion ofsaid cover inserted through the transverse slot of said open moldmember.

References Cited The following references, cited by the Examiner, are ofrecord in the patented le of this patent or the original patent.

UNITED STATES PATENTS 2,996,762 8/1961 McCormick 425-117 X 3,411,18511/1968 Pickett 425-117 3,456,300 7/1969 Pickett 425-117 J HOWARD FLlNT,JR., Primary Examiner U.S. C1. X.R.

